Ca2+ influx and phosphoinositide signalling are essential for the establishment and maintenance of cell polarity in monospores from the red alga Porphyra yezoensis.

Li L, Saga N, Mikami K - J. Exp. Bot. (2009)

Bottom Line:
The results indicate that the inhibition of the establishment of cell polarity, as judged by the ability of F-actin to localize asymmetrically, cell wall synthesis, and development into germlings, occurred when monospores were treated with inhibitors of the Ca2+ permeable channel, phospholipase C (PLC), diacylglycerol kinase, and inositol-1,4,5-trisphosphate receptor.Moreover, it was also found that light triggered the establishment of cell polarity via photosynthetic activity but not its direction, indicating that the Ca2+ influx and PLC activation required for the establishment of cell polarity are light dependent.Taken together, these findings suggest that there is functional diversity between the PLC and PLD signalling systems in terms of the formation of cell polarity; the former being critical for the light-dependent establishment of cell polarity and the latter playing a role in the maintenance of established cell polarity.

ABSTRACTThe asymmetrical distribution of F-actin directed by cell polarity has been observed during the migration of monospores from the red alga Porphyra yezoensis. The significance of Ca2+ influx and phosphoinositide signalling during the formation of cell polarity in migrating monospores was analysed pharmacologically. The results indicate that the inhibition of the establishment of cell polarity, as judged by the ability of F-actin to localize asymmetrically, cell wall synthesis, and development into germlings, occurred when monospores were treated with inhibitors of the Ca2+ permeable channel, phospholipase C (PLC), diacylglycerol kinase, and inositol-1,4,5-trisphosphate receptor. Moreover, it was also found that light triggered the establishment of cell polarity via photosynthetic activity but not its direction, indicating that the Ca2+ influx and PLC activation required for the establishment of cell polarity are light dependent. By contrast, inhibition of phospholipase D (PLD) prevented the migration of monospores but not the asymmetrical localization of F-actin. Taken together, these findings suggest that there is functional diversity between the PLC and PLD signalling systems in terms of the formation of cell polarity; the former being critical for the light-dependent establishment of cell polarity and the latter playing a role in the maintenance of established cell polarity.

fig2: Involvement of PLC activity in the establishment of cell polarity in monospores. (A) Effects of PLC inhibitor on the motility and development of monospores. Freshly released monospores incubated with an increasing concentration of U73122 from 0.1 nM to 1 μM and 1 μM U73343 for 3 h (a) and 24 h (b). Columns and vertical bars represent the mean and SD, respectively (n=3). (B) Effects of PLC inhibitor on the F-actin organization and renascent cell wall synthesis. Freshly released monospores incubated with 1 μM U73122 (a, c, e) and 1 μM U73343 (b, d, f) for 3 h (a, b), 8 h (c, d), and 24 h (e, f). The organization of F-actin in U73122 and U73343 treated monospores are indicated in (a, c) and (b, d). Renascent cell wall synthesis in U73122 and U73343 treated monospores are indicated in (e) and (f). Arrow in (b) indicates the direction of migration of the monospore. Left and right photographs in each panel show bright-field and fluorescent images, respectively. Scale bars=5 μm.

Mentions:
Since the activity of PLC depends on [Ca2+]cyt in eukaryotic cells (Staxén et al., 1999; Pan et al., 2005), the effects of U73122, a specific inhibitor of PLC (Smith et al., 1990), were tested on motility and germling formation in freshly released monospores. In the presence of U73122, monospores did not start moving or develop into germlings at concentrations ranging from 10 nM to 1 μM (Fig. 2Aa, b), concentrations that are much lower than those used in studies of other higher plants and animals (10–100 μM). Figure 2B shows that F-actin distributed symmetrically after incubation with 1 μM U73122 for 3 h and 8 h (Fig. 2Ba, c), and, moreover, no cell wall was observed in these cells after 24 h incubation (Fig. 2Be). The effects of U73122 were not reversible at either 1 μM or 0.1 μM after washing (data not shown). By contrast, the inactive analogue U73343 showed no effect on motility or further development of the freshly released monospores at 1 μM (Fig. 2Aa, b). In this case, monospores started to move and form germlings normally, with polarized F-actin observed at the leading edge of the migrating monospores (Fig. 2Bb) and at the bottom of 1-celled germlings (Fig. 2Bd). Moreover, a thick renascent cell wall was found in germlings after 24 h incubation with U73343 (Fig. 2Bf). These results indicate that PLC is involved in the establishment of cell polarity to direct the asymmetrical localization of F-action and the subsequent migration of monospores.

fig2: Involvement of PLC activity in the establishment of cell polarity in monospores. (A) Effects of PLC inhibitor on the motility and development of monospores. Freshly released monospores incubated with an increasing concentration of U73122 from 0.1 nM to 1 μM and 1 μM U73343 for 3 h (a) and 24 h (b). Columns and vertical bars represent the mean and SD, respectively (n=3). (B) Effects of PLC inhibitor on the F-actin organization and renascent cell wall synthesis. Freshly released monospores incubated with 1 μM U73122 (a, c, e) and 1 μM U73343 (b, d, f) for 3 h (a, b), 8 h (c, d), and 24 h (e, f). The organization of F-actin in U73122 and U73343 treated monospores are indicated in (a, c) and (b, d). Renascent cell wall synthesis in U73122 and U73343 treated monospores are indicated in (e) and (f). Arrow in (b) indicates the direction of migration of the monospore. Left and right photographs in each panel show bright-field and fluorescent images, respectively. Scale bars=5 μm.

Mentions:
Since the activity of PLC depends on [Ca2+]cyt in eukaryotic cells (Staxén et al., 1999; Pan et al., 2005), the effects of U73122, a specific inhibitor of PLC (Smith et al., 1990), were tested on motility and germling formation in freshly released monospores. In the presence of U73122, monospores did not start moving or develop into germlings at concentrations ranging from 10 nM to 1 μM (Fig. 2Aa, b), concentrations that are much lower than those used in studies of other higher plants and animals (10–100 μM). Figure 2B shows that F-actin distributed symmetrically after incubation with 1 μM U73122 for 3 h and 8 h (Fig. 2Ba, c), and, moreover, no cell wall was observed in these cells after 24 h incubation (Fig. 2Be). The effects of U73122 were not reversible at either 1 μM or 0.1 μM after washing (data not shown). By contrast, the inactive analogue U73343 showed no effect on motility or further development of the freshly released monospores at 1 μM (Fig. 2Aa, b). In this case, monospores started to move and form germlings normally, with polarized F-actin observed at the leading edge of the migrating monospores (Fig. 2Bb) and at the bottom of 1-celled germlings (Fig. 2Bd). Moreover, a thick renascent cell wall was found in germlings after 24 h incubation with U73343 (Fig. 2Bf). These results indicate that PLC is involved in the establishment of cell polarity to direct the asymmetrical localization of F-action and the subsequent migration of monospores.

Bottom Line:
The results indicate that the inhibition of the establishment of cell polarity, as judged by the ability of F-actin to localize asymmetrically, cell wall synthesis, and development into germlings, occurred when monospores were treated with inhibitors of the Ca2+ permeable channel, phospholipase C (PLC), diacylglycerol kinase, and inositol-1,4,5-trisphosphate receptor.Moreover, it was also found that light triggered the establishment of cell polarity via photosynthetic activity but not its direction, indicating that the Ca2+ influx and PLC activation required for the establishment of cell polarity are light dependent.Taken together, these findings suggest that there is functional diversity between the PLC and PLD signalling systems in terms of the formation of cell polarity; the former being critical for the light-dependent establishment of cell polarity and the latter playing a role in the maintenance of established cell polarity.

ABSTRACTThe asymmetrical distribution of F-actin directed by cell polarity has been observed during the migration of monospores from the red alga Porphyra yezoensis. The significance of Ca2+ influx and phosphoinositide signalling during the formation of cell polarity in migrating monospores was analysed pharmacologically. The results indicate that the inhibition of the establishment of cell polarity, as judged by the ability of F-actin to localize asymmetrically, cell wall synthesis, and development into germlings, occurred when monospores were treated with inhibitors of the Ca2+ permeable channel, phospholipase C (PLC), diacylglycerol kinase, and inositol-1,4,5-trisphosphate receptor. Moreover, it was also found that light triggered the establishment of cell polarity via photosynthetic activity but not its direction, indicating that the Ca2+ influx and PLC activation required for the establishment of cell polarity are light dependent. By contrast, inhibition of phospholipase D (PLD) prevented the migration of monospores but not the asymmetrical localization of F-actin. Taken together, these findings suggest that there is functional diversity between the PLC and PLD signalling systems in terms of the formation of cell polarity; the former being critical for the light-dependent establishment of cell polarity and the latter playing a role in the maintenance of established cell polarity.